Electroporated GLUT4-7myc-GFP detects in vivo glucose transporter 4 translocation in skeletal muscle without discernible changes in GFP patterns
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Electroporated GLUT4-7myc-GFP detects in vivo glucose transporter 4 translocation in skeletal muscle without discernible changes in GFP patterns. / Knudsen, Jonas Roland; Henriquez-Olguín, Carlos; Li, Zhencheng; Jensen, Thomas Elbenhardt.
In: Experimental Physiology, Vol. 104, No. 5, 2019, p. 704-714.Research output: Contribution to journal › Journal article › Research › peer-review
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T1 - Electroporated GLUT4-7myc-GFP detects in vivo glucose transporter 4 translocation in skeletal muscle without discernible changes in GFP patterns
AU - Knudsen, Jonas Roland
AU - Henriquez-Olguín, Carlos
AU - Li, Zhencheng
AU - Jensen, Thomas Elbenhardt
N1 - CURIS 2019 NEXS 098
PY - 2019
Y1 - 2019
N2 - Insulin and exercise lead to translocation of the glucose transporter 4 (GLUT4) to the surface membrane of skeletal muscle fibres. This process is pivotal for facilitating glucose uptake into skeletal muscle. To study this, a robust assay to directly measure the translocation of GLUT4 in adult skeletal muscle is needed. Here, we aimed to validate a simple GLUT4 translocation assay using a genetically encoded biosensor in mouse skeletal muscle. We transfected GLUT4-7myc-GFP into mouse muscle to study live GLUT4 movement and to evaluate GLUT4 insertion in the muscle surface membrane following in vivo running exercise and pharmacological activation of AMP activated protein kinase (AMPK). Transfection led to expression of GLUT4-7myc-GFP that were dynamic in live flexor digitorum brevis fibres and which, upon insulin stimulation, exposed the myc-epitope extracellularly. Running exercise, as well as AMPK-activation by 5-Aminoimidazole-4-carboxamide ribonucleotide, induced ∼125% and ∼100% increase in extracellularly exposure of GLUT4 in the surface membrane of tibialis anterior muscle. Interestingly, the clear increase in surface-exposed GLUT4 content by insulin, exercise or AMPK activation was not accompanied by any discernible reorganization of the GLUT4-GFP signal. In conclusion, we provide a detailed description of an easy to use translocation assay to study GLUT4 accumulation at the surface membrane by exercise and exercise-mimicking stimuli. Notably, our analyses revealed that increased GLUT4 surface membrane accumulation was not accompanied by a discernible change in the GLUT4 localization pattern.
AB - Insulin and exercise lead to translocation of the glucose transporter 4 (GLUT4) to the surface membrane of skeletal muscle fibres. This process is pivotal for facilitating glucose uptake into skeletal muscle. To study this, a robust assay to directly measure the translocation of GLUT4 in adult skeletal muscle is needed. Here, we aimed to validate a simple GLUT4 translocation assay using a genetically encoded biosensor in mouse skeletal muscle. We transfected GLUT4-7myc-GFP into mouse muscle to study live GLUT4 movement and to evaluate GLUT4 insertion in the muscle surface membrane following in vivo running exercise and pharmacological activation of AMP activated protein kinase (AMPK). Transfection led to expression of GLUT4-7myc-GFP that were dynamic in live flexor digitorum brevis fibres and which, upon insulin stimulation, exposed the myc-epitope extracellularly. Running exercise, as well as AMPK-activation by 5-Aminoimidazole-4-carboxamide ribonucleotide, induced ∼125% and ∼100% increase in extracellularly exposure of GLUT4 in the surface membrane of tibialis anterior muscle. Interestingly, the clear increase in surface-exposed GLUT4 content by insulin, exercise or AMPK activation was not accompanied by any discernible reorganization of the GLUT4-GFP signal. In conclusion, we provide a detailed description of an easy to use translocation assay to study GLUT4 accumulation at the surface membrane by exercise and exercise-mimicking stimuli. Notably, our analyses revealed that increased GLUT4 surface membrane accumulation was not accompanied by a discernible change in the GLUT4 localization pattern.
KW - Faculty of Science
KW - Skeletal muscle
KW - Exercise
KW - Glucose transport
U2 - 10.1113/EP087545
DO - 10.1113/EP087545
M3 - Journal article
C2 - 30710396
VL - 104
SP - 704
EP - 714
JO - Experimental Physiology
JF - Experimental Physiology
SN - 0958-0670
IS - 5
ER -
ID: 212911074